Tetraalkylammonium-based quaternary ammonium ([R4N+][DHP−]; R = Ethyl, Propyl, Butyl) salts were synthesized and subjected to in-depth analysis using NMR, FTIR and TG analysis. Subsequent investigations involved examining their impact on the aggregation patterns of the cationic surfactant Cetyltrimethylammonium bromide (CTAB) through the use of conductometry, fluorescence measurements, and UV-Visible spectroscopy. To gain a deeper understanding of the colloidal behavior of CTAB, the critical micelle concentration (cmc) of the surfactant was determined, both in the absence and presence of the quaternary ammonium salts. The observed variations in cmc values were attributed to structural changes within the surfactant-water mixture, made by the presence of these additives. In addition, the investigations include thermodynamic analysis, with a focus on studying the dependence of the cmc on temperature. The resultant thermodynamic parameters were utilized to elucidate the contributions of [R4N+][DHP−], to the electrostatic and hydrophobic interactions involved in the formation of surfactant micellar aggregates. Furthermore, the study assessed the impact of [R4N+][DHP−], on the surfactant’s antibacterial activity by quantifying the zone of inhibition, against both gram-positive and gram-negative bacterial species.